A monoclonal antibody-producing hybridoma cell line has been raised against merozoites of Babesia divergens. This antibody is strongly reactive against merozoites in an enzyme-linked immunosorbent assay (ELISA) and is also positive in an indirect fluorescent antibody test (IFAT) on preparations of live merozoites. The antibody recognizes an antigen, of molecular weight (Mr) 50-60 K by Western blot analysis on SDS-polyacrylamide gel electrophoresis (SDS-PAGE) gels. Merozoite neutralization assays show that the antibody significantly inhibits merozoite invasion of bovine erythrocytes.
A Babesia divergens merozoite antigen was purified by affinity chromatography using a monoclonal antibody. Silver staining of SDS-PAGE gels revealed 2 bands of Mr 50-60 kDa and Mr 24-29. It is proposed that the Mr 24-29 kDa band represents the native protein and that it is complexed, perhaps as a dimer or perhaps to host protein to give the Mr 50-60 kDa band. Peptide analysis of the antigen followed by Western blotting with the monoclonal antibody revealed only an Mr 20-24 kDa band. After affinity purification the antigen was precipitated with acetone and inoculated subcutaneously, together with complete Freund's adjuvant, into gerbils. Two inoculations were given 3 weeks apart, after which the immunized animals and a group of control animals were challenged with 5 x 10(7) parasitized red cells from a virulent microaerophilus stationary phase (MASP) culture of an homologous strain of B. divergens. Parasitaemias rose rapidly in control animals and all were dead by the fifth day. Two of the vaccinated animals survived the challenge. Parasitaemias in the remaining two rose slowly but both animals succumbed on the seventh day. Western blotting of merozoites and of the purified antigen, using sera from the surviving animals showed strong reactivity with the Mr 24-29 kDa band.
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